The overall goal of this proposal is to establish a novel freezing device that will prevent cell damage associated with extracellular ice formation during cryopreservation of rodent sperm. Rodent sperm have an unusual morphology compared to other mammalian species, with an asymmetric, falciform head and a relatively very long principle piece. They are also extremely mechanically sensitive. Related to this mechanical sensitivity, mouse and rat sperm are vulnerable to damage associated with the morphology of the extracellular ice formed during cryopreservation. However, current paradigms for developing cryopreservation procedures focus on factors such as the formation of intracellular ice and the high intra/extracellular solute concentrations during freezing, rather than the mechanical sensitivity of cells or ice-cell interactions. Here we proposed to develop novel devices and methods to prevent extracellular ice induced damage to rodent during cryopreservation. Our long-term goal is to establish a rodent sperm cryopreservation approach to prevent the mechanical damage to cells associated with extracellular ice formation. To that end, a novel freezing device with semi- permeable membranes will be produced to isolate rodent sperm from extracellular ice during freezing processes. PUBLIC HEALTH RELEVANCE: The overall goal of this proposal is to establish a novel freezing device that will prevent cell damage associated with extracellular ice formation during cryopreservation of rodent sperm.